Epigenome-wide association study of alcohol consumption in N = 8161 individuals and relevance to alcohol use disorder pathophysiology: identification of the cystine/glutamate transporter SLC7A11 as a top target.

Alcohol misuse is common in many societies worldwide and is associated with extensive morbidity and mortality, often leading to alcohol use disorders (AUD) and alcohol-related end-organ damage. The underlying mechanisms contributing to the development of AUD are largely unknown; however, growing evidence suggests that alcohol consumption is strongly associated with alterations in DNA methylation. Identification of alcohol-associated methylomic variation might provide novel insights into pathophysiology and novel treatment targets for AUD. Here we performed the largest single-cohort epigenome-wide association study (EWAS) of alcohol consumption to date (N = 8161) and cross-validated findings in AUD populations with relevant endophenotypes, as well as alcohol-related animal models. Results showed 2504 CpGs significantly associated with alcohol consumption (Bonferroni p value < 6.8 × 10-8) with the five leading probes located in SLC7A11 (p = 7.75 × 10-108), JDP2 (p = 1.44 × 10-56), GAS5 (p = 2.71 × 10-47), TRA2B (p = 3.54 × 10-42), and SLC43A1 (p = 1.18 × 10-40). Genes annotated to associated CpG sites are implicated in liver and brain function, the cellular response to alcohol and alcohol-associated diseases, including hypertension and Alzheimer's disease. Two-sample Mendelian randomization confirmed the causal relationship of consumption on AUD risk (inverse variance weighted (IVW) p = 5.37 × 10-09). A methylation-based predictor of alcohol consumption was able to discriminate AUD cases in two independent cohorts (p = 6.32 × 10-38 and p = 5.41 × 10-14). The top EWAS probe cg06690548, located in the cystine/glutamate transporter SLC7A11, was replicated in an independent cohort of AUD and control participants (N = 615) and showed strong hypomethylation in AUD (p < 10-17). Decreased CpG methylation at this probe was consistently associated with clinical measures including increased heavy drinking days (p < 10-4), increased liver function enzymes (GGT (p = 1.03 × 10-21), ALT (p = 1.29 × 10-6), and AST (p = 1.97 × 10-8)) in individuals with AUD. Postmortem brain analyses documented increased SLC7A11 expression in the frontal cortex of individuals with AUD and animal models showed marked increased expression in liver, suggesting a mechanism by which alcohol leads to hypomethylation-induced overexpression of SLC7A11. Taken together, our EWAS discovery sample and subsequent validation of the top probe in AUD suggest a strong role of abnormal glutamate signaling mediated by methylomic variation in SLC7A11. Our data are intriguing given the prominent role of glutamate signaling in brain and liver and might provide an important target for therapeutic intervention.

PCSK9 and the Gut-Liver-Brain Axis: A Novel Therapeutic Target for Immune Regulation in Alcohol Use Disorder.

Alcohol use disorder (AUD) is a chronic relapsing disorder characterized by an impaired ability to control or stop alcohol intake and is associated with organ damage including alcohol-associated liver disease (ALD) and progressive neurodegeneration. The etiology of AUD is complex, but organ injury due to chronic alcohol use can be partially attributed to systemic and local inflammation along the gut-liver-brain axis. Excessive alcohol use can result in translocation of bacterial products into circulation, increased expression of pro-inflammatory cytokines, and activation of immune cells, including macrophages and/or microglia in the liver and brain. One potential mediator of this alcohol-induced inflammation is proprotein convertase subtilisin/kexin type 9 (PCSK9). PCSK9 is primarily known for its regulation of plasma low-density lipoprotein cholesterol but has more recently been shown to influence inflammatory responses in the liver and brain. In rodent and post-mortem brain studies, chronic alcohol use altered methylation of the PCSK9 gene and increased expression of PCSK9 in the liver and cerebral spinal fluid. Additionally, PCSK9 inhibition in a rat model of ALD attenuated liver inflammation and steatosis. PCSK9 may play an important role in alcohol-induced pathologies along the gut-liver-brain axis and may be a novel therapeutic target for AUD-related liver and brain inflammation.

Epigenome-wide association study and multi-tissue replication of individuals with alcohol use disorder: evidence for abnormal glucocorticoid signaling pathway gene regulation.

Alcohol use disorder (AUD) is a chronic debilitating disorder with limited treatment options and poorly defined pathophysiology. There are substantial genetic and epigenetic components; however, the underlying mechanisms contributing to AUD remain largely unknown. We conducted the largest DNA methylation epigenome-wide association study (EWAS) analyses currently available for AUD (total N = 625) and employed a top hit replication (N = 4798) using a cross-tissue/cross-phenotypic approach with the goal of identifying novel epigenetic targets relevant to AUD. Results show that a network of differentially methylated regions in glucocorticoid signaling and inflammation-related genes were associated with alcohol use behaviors. A top probe consistently associated across all cohorts was located in the long non-coding RNA growth arrest specific five gene (GAS5) (p < 10-24). GAS5 has been implicated in regulating transcriptional activity of the glucocorticoid receptor and has multiple functions related to apoptosis, immune function and various cancers. Endophenotypic analyses using peripheral cortisol levels and neuroimaging paradigms showed that methylomic variation in GAS5 network-related probes were associated with stress phenotypes. Postmortem brain analyses documented increased GAS5 expression in the amygdala of individuals with AUD. Our data suggest that alcohol use is associated with differential methylation in the glucocorticoid system that might influence stress and inflammatory reactivity and subsequently risk for AUD.

Out of breath, out of time: interactions between HIF and circadian rhythms.

Humans have internal circadian clocks that ensure that important physiological functions occur at specific times of the day. These molecular clocks are regulated at the genomic level and exist in most cells of the body. Multiple circadian resetting cues have been identified, including light, temperature, and food. Recently, oxygen has been identified as a resetting cue, and emerging science indicates that this occurs through interactions at the cellular level between the circadian transcription-translation feedback loop and the hypoxia-inducible pathway (hypoxia-inducible factor; subject of the 2019 Nobel Prize in Physiology or Medicine). This review will cover recently identified relationships between HIF and proteins of the circadian clock. Interactions between the circadian clock and hypoxia could have wide-reaching implications for human diseases, and understanding the molecular mechanisms regulating these overlapping pathways may open up new strategies for drug discovery.

Proprotein Convertase Subtilisin/Kexin Type 9 (PCSK9) in the Brain and Relevance for Neuropsychiatric Disorders.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) has long been studied in the liver due to its regulation of plasma low-density lipoprotein cholesterol (LDL-C) and its causal role in familial hypercholesterolemia. Although PCSK9 was first discovered in cerebellar neurons undergoing apoptosis, its function in the central nervous system (CNS) is less clear. PCSK9 has been shown to be involved in neuronal differentiation, LDL receptor family metabolism, apoptosis, and inflammation in the brain, but in vitro and in vivo studies offer contradictory findings. PCSK9 expression in the adult brain is low but is highly upregulated during disease states. Cerebral spinal fluid (CSF) PCSK9 concentrations are correlated with neural tube defects and neurodegenerative diseases in human patients. Epigenetic studies reveal that chronic alcohol use may modulate methylation of the PCSK9 gene and genetic studies show that patients with gain-of-function PCSK9 variants have higher LDL-C and an increased risk of ischemic stroke. Early safety studies of the PCSK9 inhibitors evolocumab and alirocumab, used to treat hypercholesterolemia, hinted that PCSK9 inhibition may negatively impact cognition but more recent, longer-term clinical trials found no adverse neurocognitive events. The purpose of this review is to elucidate the role of PCSK9 in the brain, particularly its role in disease pathogenesis.